Capture of single-stranded DNA assisted by oligonucleotide modules.
Real-time biospecific interaction analysis was employed to monitor direct capture of a hepatitis C virus (HCV) derived polymerase chain reaction (PCR) product by nucleic acid hybridization. Different formats for hybridization were used to study the interaction between a single-stranded HCV PCR product and capture oligonucleotides immobilized on a sensor chip via streptavidin-biotin chemistry. By employing a prehybridization step in solution with nonbiotin oligonucleotides complementary to the single-stranded target and adjacent to the immobilized probe, a significant capture was achieved in comparison to the low capture efficiency obtained using single immobilized probes (9-36 mer). High capture efficiencies were also observed when shorter immobilized probes were used in combination with strings of adjacently positioned prehybridized probes (i.e., modules). Interestingly, the introduction of single nucleotide gaps between prehybridized and/or immobilized probes dramatically reduced the capture efficiency. These results suggest that flexible systems for capture could be designed from libraries of short oligonucleotides (9 mers) used in module fashion, taking advantage of stacking interactions between the oligonucleotides. The potential applications of such oligonucleotide-assisted capture systems are discussed.
O'Meara
D
D
Department of Biochemistry and Biotechnology, Royal Institute of Technology (KTH), Stockholm, Sweden.
Nilsson
P
P
Nygren
P A
PA
Uhlén
M
M
Lundeberg
J
J
D017931:DNA Primers; D004277:DNA, Single-Stranded / Q000032:analysis*; D004279:DNA, Viral / Q000032:analysis*; D016679:Genome, Viral; D016174:Hepacivirus / Q000235:genetics*; D009693:Nucleic Acid Hybridization / Q000379:methods; D009841:Oligonucleotides; D016133:Polymerase Chain Reaction / Q000379:methods*; D012680:Sensitivity and Specificity; D013698:Templates, Genetic
D017931:DNA Primers; D004277:DNA, Single-Stranded; D004279:DNA, Viral; D009841:Oligonucleotides